Phosphoproteomic Analysis of Human Mesenchymal Stromal Cells during Osteogenic Differentiation Ting Lo Chia-Feng Tsai Yu-Ru V. Shih Yi-Ting Wang Sheng-Chieh Lu Ting-Yi Sung Wen-Lian Hsu Yu-Ju Chen Oscar K. Lee 10.1021/pr200868p.s004 https://acs.figshare.com/articles/dataset/Phosphoproteomic_Analysis_of_Human_Mesenchymal_Stromal_Cells_during_Osteogenic_Differentiation/2554267 Human mesenchymal stromal cells (hMSCs) are promising candidates for cell therapy and tissue regeneration. Knowledge of the molecular mechanisms governing hMSC commitment into osteoblasts is critical to the development of therapeutic applications for human bone diseases. Because protein phosphorylation plays a critical role in signaling transduction network, the purpose of this study is to elucidate the phosphoproteomic changes in hMSCs during early osteogenic lineage commitment. hMSCs cultured in osteogenic induction medium for 0, 1, 3, and 7 days were analyzed by liquid chromatography tandem mass spectrometry (LC–MS/MS). Surprisingly, we observed a dramatic loss of protein phosphorylation level after 1 day of osteogenic induction. Pathways analysis of these reduced phosphoproteins exhibited a high correlation with cell proliferation and protein synthesis pathways. During osteogenic differentiation, differentially expressed phosphoproteins demonstrated the dynamic alterations in cytoskeleton at the early stages of differentiation. The fidelity of our quantitative phosphoproteomic analyses were further confirmed by Western blot analyses, and the changes from protein expression or its phosphorylation level were distinguished. In addition, several ion channels and transcription factors with differentially expressed phosphorylation sites during osteogenic differentiation were identified and may serve as potentially unexplored transcriptional regulators of the osteogenic phenotype of hMSCs. Taken together, our results have demonstrated the dynamic changes in phosphoproteomic profiles of hMSCs during osteogenic differentiation and unraveled potential candidates mediating the osteogenic commitment of hMSCs. The findings in this study may also shed light on the development of new therapeutic targets for metabolic bone diseases such as osteoporosis and osteomalacia. 2012-02-03 00:00:00 phosphoproteomic hMSC chromatography tandem mass spectrometry bone diseases osteogenic lineage commitment Osteogenic DifferentiationHuman mesenchymal stromal cells osteogenic induction medium osteogenic differentiation protein synthesis pathways Western blot analyses LC protein phosphorylation level Human Mesenchymal Stromal Cells